Technologies for differentiating undifferentiated cells such as stem cells into desired cells are expected to be adaptable to regenerative medicine and screening of prophylactic/therapeutic drugs for various diseases.
Conventionally, there are known methods for obtaining differentiated cells or products thereof from stem cells in vitro or in vivo.
For example, a method for obtaining differentiated cells or products thereof in vitro is described in U.S. Pat. No. 8,268,621, Japanese Unexamined Patent Publication No. 2004-135625 or US Patent Publication No. 2012/0315338.
U.S. Pat. No. 8,268,621 describes a method of producing primary germ layer cells, comprising: culturing embryonic stem cells or the like in the presence of an agent that alters the epigenetic state of cell such as a demethylation agent (5-azacytidine, 5-aza-2′ deoxycytidine, etc.) or a histone deacetylase inhibitor; and culturing the embryonic stem cells in the presence of various growth factors or cytokines.
Japanese Unexamined Patent Publication No. 2004-135625 describes a method for inducing the differentiation of somatic stem cells into somatic cells, comprising co-culturing desired tissue- and organ-derived cells with the somatic stem cells. In the method disclosed in the patent document, it is necessary to prepare desired tissue- and organ-derived cells after differentiation for co-culture.
US Patent Publication No. 2012/0315338 describes a method of generating megakaryocytes by culturing hemangioblasts in a medium containing various growth factors or cytokines and a method of generating platelets by culturing the generated megakaryocytes.
A method for obtaining a differentiated cell or a product thereof in vivo is described in, for example, Japanese Unexamined Patent Publication No. 2002-171865.
Japanese Unexamined Patent Publication No. 2002-171865 describes a method for producing human organ cells, comprising: introducing human-derived organ cells or organ stem cells through a duct of an animal other than humans; transplanting the human-derived organ cells or organ stem cells into a specific site of the animal other than humans; allowing the cells to grow and differentiate; and collecting the resultant human-derived organ cells. The patent document describes that the transplanted organ cells or organ stem cells are allowed to grow to a certain level, and the use of, for example, a fluorescence-activated cell sorter (FACS) enables human cells to be separated from animal cells according to differences of surface antigens.
WO 2007/025233 describes a method of making a solid organ to be transplanted into a patient by using stem cells. The patent document describes a method for producing an organ, comprising: perfusing an organ with a cell disruption medium containing a surfactant to form a decellularized organ; and bringing the decellularized organ into contact with populations of regenerated cells such as mesenchymal stem cells (MSC), under conditions such that the regenerated cells are allowed to engraft, grow, and/or differentiate inside or on the surface of the decellularized organ.
Japanese Unexamined Patent Publication No. 2012-019690 describes a method for evaluating whether or not stem cells can differentiate into cells forming a desired tissue in vivo. In the method described in the patent document, the stem cells to be evaluated are transplanted into a desired tissue anlage of a non-human mammal, and then the tissue anlage is cultured in vitro. The possibility of differentiation of the stem cells into cells forming the tissue in vivo is determined by using the level of distribution of the cells, in the tissue anlage, derived from the transplanted stem cells as an indicator.
Although the method described in U.S. Pat. No. 8,268,621 and the method described in US Patent Publication No. 2012/0315338 describe a method for obtaining differentiated cells or products thereof by culturing stem cells in a medium containing a predetermined substance, culture conditions are complicated because it is necessary to subculture the cultured cells multiple times.
In the method described in Japanese Unexamined Patent Publication No. 2004-135625, it is necessary to prepare differentiated cells for co-culture, in order to allow stem cells to differentiate. Accordingly, in the case where it is difficult to obtain differentiated cells or to culture differentiated cells, it is not resultantly possible to allow the stem cells to differentiate into target cells.
In the method of obtaining differentiated cells from stem cells in vivo as disclosed in Japanese Unexamined Patent Publication No. 2002-171865, it is inevitable that the differentiated cells to be obtained are mixed with host cells. Consequently, the subsequent purification process becomes a burden.
In WO 2007/025233, it is necessary to perform decellularization by perfusing an organ with a cell disruption medium containing a surfactant, which makes the operation complicated.
The method described in Japanese Unexamined Patent Publication No. 2012-019690 aims at determining whether or not stem cells to be transplanted are suitable for transplantation, and it does not assume the process of collecting differentiated cells.
An object of the present invention is to provide a method for obtaining differentiated cells and/or differentiated cell products from undifferentiated cells, which is a simple operational method and enables differentiated cells to be easily obtained.